Disturbances of iron metabolism increase morbidity and mortality and are among the most common disorders affecting humans. Nearly 20% of women of reproductive age in the US are iron deficient, and iron overload is increasingly being recognized as a public health concern. Despite the prevalence and adverse health effects associated with these disorders, many questions remain regarding the molecular mechanisms of iron transport. The long-term objective of the proposed work is to enhance our understanding of iron homeostasis by investigating the regulation and function of ZIP14 and other ZIP family members. ZIP14 was first identified as a zinc transporter, but recent studies indicate that it transports iron as well. Its abundant expression in the liver suggests that it plays a role in hepatic iron deposition during iron overload, and its upregulation in liver by phlebotomy and iron deficiency suggests that it functions in iron uptake. The first aim of the proposed research will be to investigate more completely the iron-dependent regulation of Zip14. Rats and mice will be made iron deficient, iron normal, or iron loaded, and a variety of tissues will be examined for Zip14 expression and cellular localization. In the second aim, cell culture studies will be used to identify the subcellular localization of Zip14 and to investigate its role in the uptake of transferrin-bound iron, the most common pathway of iron uptake by cells. To better define the in vivo role of Zip14, the third aim will characterize the iron status of Zip14 knockout mice. Tissue-specific knockout mice will be used to test the hypotheses that Zip14 plays a role in the uptake of non-transferrin-bound iron by the liver and dietary iron by the intestine. In the fourth aim, the iron transport activity of all mammalian ZIP proteins will be systematically assessed by overexpressing the proteins and measuring the uptake of radiolabeled iron. We will also examine the effect of in vivo iron status on the expression of all 14 ZIP family members. We anticipate that information derived from the experiments with Zip14, and perhaps other ZIP proteins, will be relevant to disorders of iron metabolism. Identification of other ZIP proteins that are capable of transporting iron or are regulated by iron status will enhance our basic understanding of iron homeostasis and metal ion trafficking in general. Disturbances of iron metabolism increase morbidity and mortality and are among the most common disorders affecting humans. Despite the prevalence and adverse health effects associated with these disorders, many questions remain regarding the molecular mechanisms of iron transport. The research described in this proposal will enhance our knowledge of iron transport, which will ultimately help to identify therapeutic targets for treating disorders of iron metabolism. PUBLIC HEALTH RELEVANCE: Disturbances of iron metabolism increase morbidity and mortality and are among the most common disorders affecting humans. Despite the prevalence and adverse health effects associated with these disorders, many questions remain regarding the molecular mechanisms of iron transport. The research described in this proposal will enhance our knowledge of iron transport, which will ultimately help to identify therapeutic targets for treating disorders of iron metabolism.